A novel self-deleting retroviral vector carrying an additional sequence recognized by the viral integrase (IN)

During retroviral integration, the viral integrase recognizes the attachment (att) sequence (formed by juxtaposition of two LTRs ends) as the substrate of integration. We have developed a self-deleting Avian Leukosis and Sarcoma Viruses (ALSVs)-based retroviral vector carrying an additional copy of the att sequence, between neo and puro genes. We observed that: (i) the resulting NP3Catt vector was produced at neo and puro titers respectively smaller and higher than that of the parental vector devoid of the att sequence; (ii) 61% of NP3Catt proviruses were flanked by LTRs; most of them were deleted of internal sequences, probably during the reverse transcription step; (iii) 31% of clones were deleted of the whole 5′ part of their genome and were flanked, in 5′, by the additional att sequence and, in 3′, by an LTR. Integration of these last proviruses was often imprecise with respect to the viral ends. At total, 77% of proviruses had lost the packaging signal and were not mobilizable by a replication-competent virus and 92% had lost the selectable gene in a single round of replication. Although still to improve, the att vector could be considered as an interesting new safe retroviral vector for gene transfer experiments.